DESCRIPTION. State the application's broad, long-term objectives and specific aims,making reference to the health relatednessoftheproject.Describe concisely the researchdesignand methodsfor achievingthesegoals. Avoid summariesof past accomplishmentsand the use of the first person. This description is meant to serve as a succinct and accurate description of the proposed work when separated from the application. If the application is funded, this description, as is, will become public information. Therefore, do notinclude proprietary/confidential information. DO NOT EXCEED THE SPACE PROVIDED. Aquaporin water channels are believed to have a key role in the urinary concentrating mechanism and the pathophysiology of Nephrogenic Diabetes Insipidus (NDI). The kidney expresses at least four aquaporins: AQP1 in proximal tubule, TDLH and vasa recta; AQP2 in apical membrane of principal cells in collecting duct; and AQP3 and AQP4 in basolateral membrane of the same cells. Specific Aim 1. To define the role of aquaporins in the urinary concentrating mechanism using transgenic knockout mice. Our lab developed the first transgenic mouse models to study water channel function - AQP1 and AQP4 knockout mice. These and AQP3 null mice will be used to define quantitatively the role of aquaporins in renal water clearance. Renal function will be evaluated by urine/serum chemistries, isolated tubule and vasa recta microperfusion, and kidney micropuncture. Specific Aim 2. To characterize the cellular defect in hereditary NDI and to test a novel therapeutic strategy. Preliminary data indicate that AQP2 mutations cause NDI by heterogeneous mechanisms involving defective AQP2 water channel function, accelerated degradation, and defective intracellular processing with ERretention. Transfected cells expressing NDI-causing AQP2 mutants will be used to characterize AQP2 misfolding, degradation mechanisms, and interactions with molecular chaperones. A mouse model of NDI will be developed and used to evaluate the efficacy of chemical chaperones to correct defective AQP2 function in NDI. Specific Aim 3. To utilize novel biophysical methods to analyze specific aspects of aquaporin structure and function. Green fluorescent protein (GFP)-aquaporin chimeras will be used to define aquaporin mobility and association state in membranes, and the cell biology of AQP2 trafficking. Methods will include fluorescence photobleaching recovery, energy transfer and ratio imaging. Also, specific water/solute transporting properties (Pd, c^)of the mammalian aquaporins will be measured.